Montmorency tart cherry (Prunus cerasus L., MTC) is famous for its abundant polyphenolic content and its potential anti-inflammatory and antioxidant effects. Despite this, the metabolic processing and bioavailability of MTC polyphenols in the human body remain poorly understood. This research sought to identify the polyphenolic metabolites of MTC formulations in urine, plasma, and stool through targeted LC-MS/MS analysis. The profiling of urine metabolites was performed using a crossover design with 12 healthy adults (23 ± 3 years). Participants ingested two MTC formulations: juice (240 mL) and powder (2 × 0.5 g capsules) during separate 48-hour interventions, with a 14-day washout period in between. Urine samples were collected at six intervals following ingestion. For plasma and stool metabolite analysis, 58 participants (aged 18–50 years) were involved in a randomized, placebo-controlled 30-day intervention. Active groups received either MTC juice (2 × 240 mL/day) or freeze-dried powder (2 × 0.5 g capsules/day), while control groups were given matched placebos. Samples were collected at four intervals to evaluate systemic absorption and gut microbial metabolism. Utilizing a TSQ Altis Triple Quadrupole LC-MS/MS, more than 30 secondary metabolites were identified, including sulfated, glucuronidated, and methylated derivatives of caffeic acid, epicatechin, and chlorogenic acid. Temporal analysis indicated rapid urinary excretion of Phase II metabolites (2–8 h) and a prolonged presence of microbial catabolites like dihydroferulic acid in stool samples for up to 48 h. Plasma metabolite profiles confirmed systemic availability and differences specific to the formulation. Stool metabolites showed the excretion of primary and secondary metabolites of polyphenols out of the body. These results illustrate that MTC polyphenols undergo significant biotransformation and are distributed differently across biological matrices. The findings offer foundational insights into the kinetics and mechanisms that contribute to the health benefits of MTC in clinical populations.